This project focuses on the structure, regulation, and function of genes involved in insect immunity, which shows close parallels to innate immunity in mammals. Our work concentrates on two model dipteran insects, the fruitfly Drosophila melanogaster and the malaria vector mosquito Anopheles gambiae. Because of its powerful genetics, Drosophila is an excellent model system in which to isolate new genes involved in immunity, and to functionally characterize their protein products. Anopheles is a system of choice to decipher the innate response of vector insects to the medically important parasites they carry. Previous work using the Drosophila model has led to the identification of many immune inducible antimicrobial peptides. Interestingly, these peptides can be grouped according to their targets, with some being active against fungi, and other against Gram-negative and/or Gram-positive bacteria. It has recently been shown that the antimicrobial response of Drosophila is not non- specific but can discriminate among various classes of microorganisms, reflected in differential expression of genes encoding antibacterial and antifungal peptides following injection of distinct microorganisms. Similarly, different levels of induction of immune markers by different bacteria have been observed in cell lines derived from the mosquito A. gambiae. In addition, recent work has revealed that exposure of surface epithelia to microorganisms results in induction of an immune response in Anopheles and Drosophila. Taken together, these results raise the question of the nature, structure and function of the molecules involved in infectious non-self recognition in insects, elucidation of which constitutes the main goal of this proposal. Over the next few years, we plan to 1) characterize molecularly the factors involved in recognition of fungi, bacteria, and protozoan parasites in Drosophila and Anopheles; 2) examine the functional importance and roles of these molecules in insect systemic and local immunity; 3) identify the molecules mediating the transduction of recognition into an immune response. Abundant precedents exist testifying that evolutionary conservation between insects and mammals is such that comparative analysis of regulatory phenomena in these two groups of animals is very fruitful. To give but one example from an earlier collaboration that led to this Programme Project application, the demonstration that the TOLL receptor is involved in the antifungal immunity in Drosophila directly stimulated the identification and functional characterization of the TOLL homologue in humans. We are applying jointly with our colleagues for a Programme Project Grant because we are confident that the structural and functional comparison of molecules recognizing molecular patterns on thy surface of microorganisms in mammals and insects will provide better insight into the mechanisms and evolution of first line host defense. In addition, these studies will aid understanding of how insects react to infection, and can be refractory to transmission of medically important parasites.